种子特异表达异源DGAT1基因提高大豆种子含油量和营养品质

Seed-specific expression of heterologous gene DGAT1 increase soybean seed oil content and nutritional quality

摘要:

为提高大豆Glycine max种子含油量和营养品质，文中以二酰甘油酰基转移酶1 (Diacylglycerol acyltransferase 1，DGAT1) 基因为遗传修饰靶标。将来自高油植物斑鸠菊Vernonia galamensis L.编码DGAT1酶蛋白的cDNA克隆 VgDGAT1A在大豆种子特异超表达。连续选择获得高代 (T7) VgDGAT1A转基因大豆株系。转基因株系表型鉴定显示，在大豆种子发育中期 (30–45 DAF)，VgDGAT1A高表达，相应地DGAT酶活性是非转基因野生型和空载体转化对照的7.8倍。转基因成熟种子含油量比对照提高了5.1%，淀粉含量比对照减少2%–3%，蛋白质含量与对照无显著差异。此外，转基因大豆种子百粒重 (14.5 g) 和种子萌发率 (95.6%) 与对照亦无明显差异。种子油脂脂肪酸成分分析显示，转基因大豆种子油中抗氧化的油酸 (C18:1Δ9) 含量比对照提高8.2%，相应地易氧化的亚油酸 (C18:2Δ9,12) 和亚麻酸 (C18:3Δ9,12,15) 分别减少6%和2%。这些数据表明，种子特异超表达外源VgDGAT1A基因，打破了大豆种子含油量和蛋白质含量的负连锁，显著提高种子含油量且未导致蛋白含量降低。转基因大豆种子重量和萌发率亦未显负效应，而且种子油脂抗氧化性和营养品质得以改善。研究表明应用这一高酶活性VgDGAT1A的基因工程是提高种子含油量和改善油脂品质的一条有效途径。

Abstract:

Enhancing soybean (Glycine max) oil production is crucial to meet the market demand of vegetable oil. Diacylglycerol acyltransferase (DGAT) catalyzes the final acylation reaction of triacylglycerol (TAG) synthesis, acting as one of the rate-limiting enzymes for oil biosynthesis in plant seeds. Here, a cDNA clone VgDGAT1A encoding the DGAT1 protein was isolated from the high oil plant Vernonia galamensis. VgDGAT1A was specifically overexpressed in soybean seeds, and several high-generation transgenic lines (T7) were obtained by continuous selection. qPCR analysis showed that VgDGAT1A was highly expressed in the mid-development stage (30–45 DAF) of the transgenic seeds. Accordingly, the DGAT enzyme activity in the transgenic seeds was increased by 7.8 folds in comparison with the wild-type controls. Seed oil and starch contents were, respectively, increased by 5.1% (Dry weight) and reduced by 2%–3% in the transgenic soybeans. Importantly, protein content was not significantly different between transgenic and control seeds. Seed weight and germination rate of the transgenic lines exhibited no negative effect. Fatty acid profiling demonstrated that antioxidant oleic acid (C18:1Δ9) content in the transgenic seed oil was elevated by 8.2% compared to the control, and correspondingly, easily-oxidized linoleic acid (C18:2Δ9,12) and linolenic acid (C18:3Δ9,12,15) were decreased by 6% and 2% respectively. Taken together, seed-specific overexpression of an exogenous VgDGAT1A gene can break the negative linkage of oil and protein contents in soybean seeds, indicating that engineering of this highly-active DGAT enzyme is an effective strategy to improve oil yield and nutritional value in oilseeds.